Peptidomimetic α-Acyloxymethylketone Warheads with Six-Membered Lactam P1 Glutamine Mimic: SARS-CoV-2 3CL Protease Inhibition, Coronavirus Antiviral Activity, and <i>in Vitro</i> Biological Stability

Abstract

Recurring coronavirus outbreaks, such as the current COVID-19 pandemic, establish a necessity to develop direct-acting antivirals that can be readily administered and are active against a broad spectrum of coronaviruses. Described in this Article are novel α-acyloxymethylketone warhead peptidomimetic compounds with a six-membered lactam glutamine mimic in P1. Compounds with potent SARS-CoV-2 3CL protease and <i>in vitro</i> viral replication inhibition were identified with low cytotoxicity and good plasma and glutathione stability. Compounds <b>15e</b>, <b>15h</b>, and <b>15l</b> displayed selectivity for SARS-CoV-2 3CL protease over CatB and CatS and superior <i>in vitro</i> SARS-CoV-2 antiviral replication inhibition compared with the reported peptidomimetic inhibitors with other warheads. The cocrystallization of <b>15l</b> with SARS-CoV-2 3CL protease confirmed the formation of a covalent adduct. α-Acyloxymethylketone compounds also exhibited antiviral activity against an alphacoronavirus and non-SARS betacoronavirus strains with similar potency and a better selectivity index than remdesivir. These findings demonstrate the potential of the substituted heteroaromatic and aliphatic α-acyloxymethylketone warheads as coronavirus inhibitors, and the described results provide a basis for further optimization.